The current application is submitted in response to the notice number NOT-OD-058 entitled "Enabling RPGs to Leverage NCRR Center and Centers-like Programs". The research focus of the parent grant RO1AI0470889 is to develop a vaccine targeting human malaria. Transmission blocking vaccines (TBVs) against malaria are intended to induce immunity against the stages of the parasite that infect mosquitoes so that malaria transmission is reduced or halted. The target antigens include proteins synthesized in the gametocytes (pre-fertilization antigens, in P. falciparum: Pfs230 and Pfs48/45) and in the zygotes-ookinetes (post-fertilization antigens, in P. falciparum: Pfs25 and Pfs28) and the epitopes recognized by transmission blocking antibodies are cysteine-rich reduction-sensitive conformational in nature. Studies proposed in the parent grant were aimed at (1) identifying immunologically relevant domains in the pre- fertilization antigens, (2) optimizing the combination of pre- and post-fertilization antigens by vaccine formulation in cationic lipids and vaccine delivery by in vivo electroporation, (3) evaluating a candidate DNA vaccine by in vivo electroporation in nonhuman primates (Macaca mulatta) and testing the concept that immunity against pre-fertilization antigens can be maintained by boosting during natural infection using an Aotus model for P. falciparum infection. Moreover, the development of Pfs25 transgenic P. berghei will provide an approach for in vivo evaluation of human malaria TBV based on Pfs25, as compared to a standard in vitro membrane feeding assay. Since most vaccines go through immunogenicity and functional evaluation (wherever biologically feasible) in nonhuman primates, it is now proposed to revise the scope of the parent grant to develop transgenic nonhuman malaria parasite expressing Pfs25 which would then facilitate optimization and evaluation of Pfs25-based TBV. PUBLIC HEALTH RELEVANCE: Malaria causes more than 300 million infections worldwide. Our long term goal is to develop a vaccine to stop transmission of malaria. Research proposed in this application, in particular, will result in the development of a nonhuman primate model to test such human malaria transmission blocking vaccines.